The worldwide demand for high potency sweeteners is increasing and, with blending of different sweeteners becoming a standard practice, the demand for alternatives is expected to increase. Such sweeteners include both caloric and low-caloric sweeteners. Caloric sweeteners include sucrose, fructose, and glucose. Recently, low-calorie (or non-calorie) sweeteners have gained increased popularity. These can be used as substitutes for caloric sweeteners and are often referred to as “sugar substitutes”. Common sugar substitutes include saccharin, aspartame, and sucralose.
One such low-calorie sweetener is stevia, which is a sweetener derived from Steviol glycosides. Demand for Steviol glycosides is growing because of their non-toxic nature, their sugar-like taste profile, and their low caloric value, when used as sugar substitutes.
Stevia rebaudiana Bertoni is a perennial shrub of the Asteraceae (Compositae) family native to certain regions of South America. Its leaves have been traditionally used for hundreds of years in Paraguay and Brazil to sweeten local teas and medicines. The plant is commercially cultivated in Japan, Singapore, Taiwan, Malaysia, South Korea, China, Israel, India, Brazil, Australia and Paraguay.
The leaves of the plant contain a mixture containing diterpene glycosides in an amount ranging from about 10 to 20% of the total dry weight. These diterpene glycosides are about 150 to 450 times sweeter than sugar. Structurally, the diterpene glycosides are characterized by a single base, steviol, and differ by the presence of carbohydrate residues at positions C13(R2) and C19(R1). The structure of the steviol base is shown hereinbelow:

Table 1 illustrates the various steviol compounds with reference to the above steviol base.
TABLE 1Chemical Structures of Steviol GlycosidesR-Groups in Stevia StructureCompoundR1R2FormulaMWRebaudioside Aβ-glc-(β-glc)2-β-glc-C44H70O23967.01Rebaudioside BH(β-glc)2-β-glc-C38H60O18804.88Rebaudioside Cβ-glc-(β-glc,α-rha-)-β-glcC44H70O22951.01Rebaudioside Dβ-glc-β-glc-(β-glc)2-β-glc-C50H80O281129.15Rebaudioside Eβ-glc-β-glc-β-glc-β-glc-C44H70O23967.01Rebaudioside Fβ-glc-(β-glc,β-xyl)-β-glc-C43H68O22936.99Rebaudioside M(β-glc)2-β-glc-(β-glc)2-β-glcC56H90O331291.3Steviosideβ-glc-β-glc-β-glc-C38H60O18804.88SteviolbiosideHβ-glc-β-glc-C32H50O13642.73Rubusosideβ-glc-β-glc-C32H50O13642.73Dulcoside Aβ-glc-α-rha-β-glc-C38H60O17788.87Glc—glucose;rha—rhammose;xyl—xylose
Typically, on a dry weight or anhydrous basis, the four major steviol glycosides found in the leaves of stevia are Dulcoside A (0.3%), Rebaudioside C (0.6-1.0%), Rebaudioside A (3.8%) and Stevioside (9.1%). Other steviol glycosides identified in stevia extract include Rebaudioside B, D, E, and F, Steviolbioside and Rubusoside. Among these, only Stevioside and Rebaudioside A are available on a commercial scale. Stevioside and Rebaudioside A are the component glycosides of principal interest for their sweetening property.
Steviol glycosides can be extracted from leaves using either water or organic solvent extraction. Typically, steviol glycosides are obtained from the leaves of Stevia rebaudiana Bertoni. The leaves are extracted with hot water and the resulting aqueous extract is passed through an adsorption resin to trap and concentrate the component steviol glycosides. Generally, the resin is desorbed by washing the resin with organic solvents like methanol or ethanol to release the glycosides. Typically, the steviol product is recrystallized with a solvent such as methanol or ethanol. Typically, the steviol product is recrystallized with a solvent such as methanol. Ion-exchange resins have been used in the purification process. The final product is typically spray-dried. (See FAO JECFA Monographs 4 (2007).
Supercritical fluid extraction and steam distillation methods have also been described. Methods for the recovery of diterpene sweet glycosides from Stevia rebaudiana Bertoni using supercritical CO2, membrane technology, and water or organic solvents, such as methanol and ethanol, may also be used.
The wide use of steviol glycosides as sweeteners has been limited to date by the presence of certain undesirable taste properties, including licorice taste, bitterness, astringency, sweet aftertaste, bitter aftertaste, licorice aftertaste. The main sweetening component of stevia extract is rebaudioside A. Rebaudioside A provides the greatest degree of sweetening without the undesirable taste properties. Many of these undesirable taste properties can be minimized or eliminated by separating the steviol glycosides, particularly rebaudioside A from other rebaudioside isomers and other compounds associated with the plant extract. Such other compounds include: proteins, resins, organic acids, pigments and sesquiterpene lactones. The pigments include chlorophyll, xanthophyll and betacarotene.
Because the chemical structures of the steviol glycosides are very similar, obtaining a relatively pure form of rebaudioside A from the mixture of other isomers is a challenge. At present the published methods for purification of rebaudioside A typically require a cascade of process steps including: filtration, precipitation of undesired components, decolorization, anion and cation exchange, and multi-stage crystallization to provide a purity of 95 weight percent on a dry or anhydrous basis. Often these steps include at least four solvent changes, drying and resolving steps.
U.S. Pat. No. 6,228,996 discloses a method for purifying diterpene glycosides from plant sources wherein the plant components such as fruit, leaves, branches, and bark, etc., are extracted to obtain a liquid extract. The liquid extract is admixed with a saturated solution containing at least one metallic ion having a valence of 2 or 3 (preferably, Al+++), and the resulting admixture is contacted with a resin to adsorb the diterpene glycosides of interest. The diterpene glycosides are desorbed from the resin by washing the resin with an alcohol solution to obtain an alcohol solution containing the diterpene glycosides. The alcohol solution is subsequently dried to provide a dry composition containing the diterpene glycosides.
U.S. Pat. No. 9,169,285 discloses methods for purifying steviol glycosides which include (a) passing a solution of steviol glycosides through a multi-column system including a plurality of columns packed with an adsorbent resin to provide at least one column having adsorbed steviol glycosides; (b) eluting the adsorbed fractions from the at least one column having adsorbed steviol glycosides using a desorbent being a solution comprising alcohol and water to provide an eluted alcoholic solution with high steviol glycoside content. Further processing steps include ion-exchange and decolorizing the eluted solution before solvent removal and drying steps to obtain a solid steviol glycoside product.
Over forty years ago, a new process was developed specifically for large scale industrial purifications. U.S. Pat. No. 2,985,589 disclosed a chromatography system involving a separation tower divided into a number of individual separation beds. These beds are connected in series, and the outlet at the bottom most bed is connected to a pump that returned flow in a continuous loop to the upper most bed. The inlet apparatus for each bed has a port connected to a downward flowing conduit. The conduits terminate in fittings attached to a rotary valve designed to control both ingress and egress of liquids into or from the inlets to each individual bed. The system is called Simulated Moving Bed (SMB) chromatography because the beds appear to be moving in a direction countercurrent to the direction of flow. There are hundreds of adsorbents which have been used for simulated moving bed systems, some of which include resins, zeolites, alumina, and silica.
Simulated Moving Bed (SMB) technology represents a variation on the principles of high performance liquid chromatography. SMB can be used to separate particles and/or chemical compounds that would be difficult or impossible to separate by any other means. Furthermore, SMB technology represents a continuous process which provides a significant economic and efficiency advantages in manufacturing operations compared to batch typical batch separation methods including crystallization and stepwise chromatographic separations.
Conventional methods for the purification of Steviol glycoside extracts are associated almost exclusively with the use of organic solvents, such as methanol, ethanol or ether. Typically, such methods require that the Steviol glycosides be initially absorbed on a resin, followed by elution of the adsorbed Steviol glycosides with an organic solvent. Thus concentrated, the resulting organic Steviol glycoside solutions are evaporated and further treated with an alcohol such as methanol or ethanol in a crystallization step to provide a purified, crystallized steviol glycoside product. To satisfy the growing demand for the stevia based sweeteners which meet commercial food quality requirements, there is a need for an efficient extraction process that can be carried out to produce the main sweetening components without the use of organic solvents. The potential for even small amounts of organic solvents remaining in the purified stevia glycoside product can be deleterious to human health.